Alcohol abuse has become a problem of epidemic proportions in industrialized countries. The accepted medical treatment includes counselling and behavior modification. However, other than asking for a direct admission from a patient, few tests exist to determine whether that patient is an alcoholic or a chronic alcohol user.
Many of the tests which have been developed rely upon biochemical markers which reflect changes in particular liver enzymes which indicate hepatic damage or abnormal hematological tests which are impaired by prolonged elevation in blood alcohol levels. The value of these tests is measured by their sensitivity (i.e., the percent of alcoholics showing a positive test) and specificity (i.e., the percent of non-alcoholics showing a negative test). The sensitivity and specificity of these tests is not high because other forms of liver injury or abnormalities other than alcoholism, influence the biological markers. Because of the influence of these other factors, tests based upon a marker directly derived from a metabolite of ethanol, such as acetaldehyde, which is not influenced by conditions unrelated to alcohol abuse would have advantages.
Acetaldehyde (AcH), the primary metabolite of ethanol, has been known to bind covalently to form stable adducts with many proteins including hemoglobin (Stevens, V. J. et al., J. Clin. Invest. 67:361-69, 1981) and it has been proposed to use these adducts as a biological marker for alcoholism and chronic alcohol use. Acetaldehyde adducts can be detected in blood from volunteers who have consumed alcohol (Niemela, O, et al., Alcoholism: Clin. Exp. Res. 14:838-41, 1990) and in the serum of alcoholic patients (Lin, RC, et al., Alcoholism: Clin. Exp. Res. 14:438-43, 1990) using various antibodies developed against protein-AcH adducts in enzyme-linked immunosorbent assays (ELISA). Unfortunately, these assays lack specificity owing to difficulties in the selection of specific Ach-protein adducts for development of antibody formation.
In 1984, Homaidan reported that acetaldehyde-hemoglobin adducts were an unreliable marker of alcohol abuse (Homaidan, F. et al., Clin. Chem. 30:480-82, 1984). Homaidan was unable to detect differences between alcoholics and social drinkers in the amounts of so-called "fast hemoglobins" (variants of HbA.sub.0) produced by acetylation by acetaldehyde or glycosylation by sugars when measured by cation-exchange chromatography or by agar gel electrophoresis.
The separation of hemoglobin fractions by high pressure liquid chromatography (HPLC), offers markedly improved resolution of hemoglobin A variants when compared to previous methods of iso-electric focusing and ion-exchange chromatography. Huisman, T. H. J. et al., J. Lab. Clin. Med. 102:163-173, 1983, may represent the earliest detection by HPLC of the Hb A.sub.1-AcH complex as an unidentified minor Hb which cochromatographed with Hb A.sub.1c in the blood of alcoholics.
Sillanaukee and Koivula (Alcoholism: Clin. Exp. Res. 4:842-46, 1990) found that AcH induces an increase in Hb A.sub.1-AcH which is one of the faster eluting hemoglobin A variants when separated by HPLC. They proposed its use as a diagnostic marker in alcoholism (as a ratio of HB A.sub.1-AcH /HB A.sub.1c) and as an indicator of heavy drinking (Sillanaukee, P. et al., Alcohol & Alcoholism 26:519-25, 1991 and Sillanaukee et al., J. Lab. Clin. Med. 120:42-47, 1992). However, difficulties have been encountered in using the Hb A.sub.1-AcH adduct as a biological indicator of alcohol abuse. The method used by Sillanaukee, inadequately resolves the Hb A.sub.1-AcH peak from the contiguous Hb A.sub.1c complex. This seriously compromises the ability of the analysis to diagnose chronic alcohol usage. The Hb A.sub.1c complex is associated with a number of nonalcoholic conditions including diabetes and, hence, is elevated in many non-heavy drinking subjects such that if the Hb A.sub.1-AcH peak does not adequately resolve from the Hb A.sub.1-Ach complex, the Hb A.sub.1-AcH peak is dwarfed by the much larger the contiguous Hb A.sub.1c peak and integration of the Hb A.sub.1-AcH peak is prevented.